A series of studies will be undertaken to elucidate the molecular mechanisms of the decreases in activity of the ketoglutarate dehydrogenase complex (KGDHC) and transketolase in Alzheimer brain, and specifically to rule out (or in) the existence of genetic abnormalities in one or more of these peptides. Stability of these enzymes in post mortem human brain will be tested by comparing their activities in autopsy and neurosurgical waste tissue. Immunochemical comparisons of the individual peptides of KGDHC and transketolase will be by ELISAs (and for KGDHC by Western blots; such blots have already been done for transketolase). In vitro translation assays will test whether or not the mRNAs coding for these peptides are functionally normal in Alzheimer tissues, using normal human peptides as markers. In vitro translation assays in the presence of appropriate subcellular fractions will test whether post-translational processing is normal (using mitochondria for KGDHC and a cytosolic fraction for transketolase). If abnormalities are found in a particular peptide or peptides, molecular genetic studies will be carried out and will concentrate on that peptide(s). They will include (a) isolating cDNA probes from a human gt11 phage library; (b) characterising these cDNAs by restriction and sequencing analyses; (c) characterization of the relevant mRNAs by Northern blots, S1- nuclease mapping, and partial sequencing with synthetic primer; (d) localization of the gene(s) on human chromosomes; (e) RFLP linkage analyses to test the relation of the genetic to the clinical abnormalities; (f) ultimately cloning and sequencing of the normal and Alzheimer gene(s). The immunochemical studies will use primarily brain, with testing for any abnormalities observed also in fibroblasts. The molecular genetic studies including in vitro translation assays will use primarily cultured cells, to avoid potential agonal and post- mortem artifacts.